Association of human papillomavirus type 16 long control region mutation and cervical cancer

Abstract

BackgroundThe variation of human papillomavirus (HPV) genes or HPV variants demonstrates different risks of cervical cancer. Mutation in the long control region (LCR) at YY1-motifs is one of the mechanisms for enhancing viral oncogene expression during the course of cancer cell progression. In Thai women, cervical cancers are almost always associated with HPV16 variant sub-lineage Asian (HPV16As); however, the mechanism involved remains elusive. The aim of this study was to understand further the oncogenic potential of HPV16As.MethodsA total of 82 HPV16-positive specimens from Thai women were selected from formalin-fixed paraffin-embedded cervical tissues, and the full length E6 gene of each specimen was amplified and sequenced. LCRs of the HPV16As-positive cases were amplified and sequenced to analyze their polymorphisms. Transcriptional activities of the HPV16As LCRs were then compared with sub-lineage European (EUR), sub-lineage Asian-American 1 (AA1) and HPV16 prototype by insertion of the LCRs into the pGL3-Basic vector.ResultsThe HPV16 DNA sequences were classified as HPV16 prototype (18.3%), Asian (As, 61%), Asian American-1 (AA1, 8.5%), European (EUR, 7.3%), Asian African-2 (AFR2, 3.7%) and Java-135C (J135C, 1.2%). The prevalence of HPV16As was 30% in low-grade squamous intraepithelial lesion (LSIL), while that in high-grade squamous intraepithelial lesion (HSIL) and squamous cell cervical carcinoma (SCC) were 63.9% and 66.7%, respectively, which demonstrates a significant association of HPV16As with the disease severity. LCR polymorphisms from 43 HPV16As positive cases were analyzed by PCR-sequencing. Thirty-eight nucleotide variation positions spanned nucleotide positions 7157–82. Ten new mutations found in the HPV16As LCRs were located predominantly at the enhancer and proximal to the 3’-end of the early promoter. The LCRs of the common HPV16As, EUR and AA1 showed 5, 13 and 23-fold higher activity than the HPV16 prototype LCR, while those of the new nucleotide variations of As showed 19 (As-sv1) and 30 (As-sv14) -fold higher activity than the HPV16 prototype.ConclusionsHPV16As DNA sequence variation, especially at the proximal to early promoter in the LCR, enhances transcriptional activity. This could be one of the possible mechanisms for HPV16As-associated cervical cancer development.

Physical Status and Variant Analysis of Human Papillomavirus 16 in Women from Shanghai

“…In our study, LCR variants were more frequently found in the EA lineage, and new mutations were only found in the EA lineage (table 4). It has been reported that disruption or deletions in the transcription factor YY1 of the viral LCR can lead to E6/E7 overexpression in primary or metastasizing tumors carrying intact E2 [48]. Among the 25 variations that we identified, 14 (52%) were within these binding sites.…”

Polymorphisms and Functional Analysis of the Intact Human Papillomavirus16 E2 Gene

“…Veress et al reported that the transcriptional activity of the HPV16 LCR variants (HPV16 AA variant) in C33A cells had higher transcriptional activity than the HPV16 LCR prototype (Veress et al, 1999). In addition, the HPV16 LCR with mutation showed increased promoter activity higher than the wild type (Chen et al, 1997;Pientong et al, 2013).…”

Activity of Andrographolide and Its Derivatives on HPV16 Pseudovirus Infection and Viral Oncogene Expression in Cervical Carcinoma Cells

“…The LCRs of the HPV16 prototype and HPV16 As lineage have different nucleotide sequences as previously reported (20). The green fluorescent protein expression vector (pEGFP-N1) was used as a marker of transfection.…”

“…C33A cells were grown in 24-well plates in DMEM with 10% FBS without antibiotics and transfected with the pGL3 reporter vector containing the LCR of HPV16 prototype or As lineage (20) and the pEGFP-N1 vector, which was the cotransfection marker, using Lipofectamine 2000. The transfected cells were treated with compounds at subcytotoxic concentrations and incubated at 37 C. All transfection assays were performed in triplicate.…”